The Bowman-Brik trypsin and chymotrypsin inhibitor (BBI), present in legume seeds, may represent a nutritionally relevant anticarcinogen. This anticarcinogenic action has been observed with pure BBI suppressing x-ray induced malignant transformation in vitro. Recently, diets supplemented with crude extracts enriched for BBI were reported to suppress dimethylhydrazine-induced colon tumors in mice. In vitro transformation studies using BBI-derivatives have also proven the chymotrypsin-inhibitory activity is responsible for supressing transformation. Studies described in this proposal aim to do the following: 1) To isolate and characterize the chymotrypsin-like enzyme from normal and transformed C3H/10T 1/2 cells. This will be accomplished using ion exchange chromatography, BBI-sepharose affinity chromatography and HPLC. 2) To quantitate levels of the BBI-inhibitable enzyme(s) in normal, "initiated" and transformed cells using a 3H-DFP protection assay coupled with HPLC. 3) To assess the cellular site of action using fluorescent BBI and fluorescent microscopy. Receptor mediated uptake will be determined using various pretreatment procedures (i.e., temperature shifts; metabolic antagonists). The long-term objective is to understand how normal cellular function is affected by the BBI sensitive enzyme(s) which presumeably hydrolyze particular protein substrates resulting in malignant cell transformation. In a larger context, the aim of this proposal is also to further elucidate the mechanism whereby a dietary protein can be a cancer preventive.